Engineer bifunctional antibacterial enzymes for treatment of S. aureus infections

设计双功能抗菌酶来治疗金黄色葡萄球菌感染

• 批准号: 9301389
• 负责人: Karl E Griswold
• 金额: $ 16.2万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-20 至 2019-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9301389
• 关键词: Address; Algorithms; Anti-Bacterial Agents; Antibiotic Resistance; Antibiotics; Antibodies; Antigen-Antibody Complex; Bacteria; Bacterial Infections; Bacteriophages; Binding; Biological Response Modifier Therapy; Bispecific Antibodies; Blood; Blood Circulation; Catalytic Domain; Cause of Death; Cell Wall; Centers for Disease Control and Prevention (U.S.); Cessation of life; Chimera organism; Chimeric Proteins; Clinic; Clinical; Communities; Cytolysis; Dangerousness; Development; Dose; Drug Costs; Drug resistance; Drug-sensitive; Economics; Endopeptidases; Engineering; Enzymes; Exhibits; Eye Infections; Fc Immunoglobulins; Fc domain; Filtration; Frequencies; Genes; Genus staphylococcus; Half-Life; Healthcare Systems; Hospitalization; Hospitals; Human; Hydrolase; Immune response; Immunoglobulin G; Immunologic Surveillance; In Vitro; Incidence; Infection; Kidney; Lung; Lysostaphin; LytA enzyme; Medical; Methicillin Resistance; Molecular; Multi-Drug Resistance; Mutation; Patients; Peptidoglycan; Performance; Phenotype; Prophages; Protein Engineering; Proteins; Radial; Recombinants; Recording of previous events; Recycling; Resistance; Resistance development; Risk; Skin; Staphylococcus aureus; Stream; T-Lymphocyte Epitopes; Technology; Testing; Therapeutic; Toxic effect; Treatment outcome; Ursidae Family; Variant; antibody engineering; bacterial resistance; bactericide; bacteriocin; base; chemotherapy; clinical translation; combat; cost; dalton; design and construction; drug development; drug discovery; drug resistant bacteria; endolysin; enzyme therapy; experience; glomerular filtration; immunogenic; immunogenicity; improved; innovation; killings; lysin; methicillin resistant Staphylococcus aureus; neonatal Fc receptor; next generation; nonhuman primate; pathogen; resistant strain; scaffold; small molecule; synergism; therapeutic candidate

Abstract: Antibiotic resistance complicates the majority of Staphylococcus aureus (S. aureus) infections. A full two thirds of hospital-associated S. aureus infections and ~50% of those acquired in the community are now methicillin-resistant (MRSA). MRSA causes >450,000 infections in the US each year, and it is responsible for half of all deaths caused by drug-resistant bacteria. The high incidence of multi-drug resistance in S. aureus and other bacteria underscores the need for next-generation antibiotics capable of combating these dangerous pathogens. An increasingly compelling therapeutic strategy leverages recombinant enzymes, such as Staphylococcus simulans lysostaphin (LST), which degrade cell wall peptidoglycan causing bacterial lysis and death. LST is a highly potent anti-staphylococcal agent with proven efficacy against both drug-sensitive and drug-resistant strains of S. aureus. While LST holds great potential for combatting dangerous S. aureus infections, its utility as a systemically administered treatment is constrained by specific limitations. First, as a small protein of 26,942 daltons, LST is rapidly cleared from the blood stream by renal filtration. In the clinic, this fact might necessitate frequent, high dosing to achieve complete bacterial clearance. Providing the option to use lower doses and less frequent administration would reduce costs, ease patient burden, and improve treatment outcomes. Second, LST is subject to development of S. aureus resistance by virtue of mutations in the femA gene, which alters the specific peptidoglycan bond targeted by the enzyme. Synergistic 2-agent treatments have been shown to mitigate the risk of LST resistance. We propose here to construct a modular bifunctional lysin platform based on fusions with immunoglobulin Fc domains. The Fc domain is a natural bivalent display scaffold, and we aim to leverage validated knob and hole bispecific Fc engineering strategies to create heterobifunctional Fc-lysin chimeras. Specifically, we will fuse the LST catalytic and cell wall binding domains to one chain of a heterodimeric knob and hole Fc, and we will fuse the SA2 prophage endopeptidase domain to the second chain. The heterologous pairing of the two Fc chains will create a single molecular entity that integrates two complementary cell wall hydrolases known to exert anti-S. aureus synergy. The bifunctional Fc-lysin's two pronged attack on S. aureus cell walls should minimize acquired resistance. Additionally, the Fc domain will serve to extend the bifunctional lysin's circulation half-life by (i) increasing the molecule's size beyond the limit for first-pass glomerular filtration in the kidney, and (ii) engaging the neonatal Fc receptor (FcRn), which actively recycles IgG antibodies and promotes their exceptionally long half-lives. As a whole, this project seeks to develop a modular platform for engineering high performance antibacterial enzymes that capitalize on intramolecular synergy to kill drug-resistant bacterial pathogens.

摘要：抗生素耐药性使大多数金黄色葡萄球菌（S. aureus）感染变得复杂。一个完整的 目前，三分之二的医院相关金黄色葡萄球菌感染以及大约 50% 的社区感染 耐甲氧西林 (MRSA)。 MRSA 每年在美国造成超过 450,000 例感染，它负责 一半的死亡是由耐药细菌引起的。金黄色葡萄球菌多重耐药发生率高 和其他细菌强调需要能够对抗这些危险的下一代抗生素 病原体。越来越引人注目的治疗策略利用重组酶，例如 模拟葡萄球菌溶葡萄球菌素 (LST)，可降解细胞壁肽聚糖，导致细菌裂解和 死亡。 LST 是一种高效的抗葡萄球菌剂，已被证明对药物敏感和 金黄色葡萄球菌的耐药菌株。虽然 LST 在对抗危险的金黄色葡萄球菌方面具有巨大潜力 感染，其作为全身治疗的效用受到特定限制的限制。首先，作为一个 LST 是 26,942 道尔顿的小蛋白质，可通过肾过滤从血流中快速清除。在诊所里，这 事实上，可能需要频繁、高剂量的给药才能实现彻底的细菌清除。提供选项 使用较低剂量和较少给药频率将降低成本、减轻患者负担并改善 治疗结果。其次，LST 会因突变而产生金黄色葡萄球菌耐药性。 femA 基因，它改变酶靶向的特定肽聚糖键。协同2剂 治疗已被证明可以降低 LST 耐药的风险。我们在这里建议构建一个模块化的 基于与免疫球蛋白 Fc 结构域融合的双功能溶素平台。 Fc 结构域是天然的 二价展示支架，我们的目标是利用经过验证的旋钮和孔双特异性 Fc 工程策略 创建异双功能 Fc-溶素嵌合体。具体来说，我们将融合 LST 催化和细胞壁结合 域到异二聚体旋钮和孔 Fc 的一条链上，我们将融合 SA2 原噬菌体内肽酶 域到第二条链。两条 Fc 链的异源配对将创建一个分子实体 它整合了两种已知可发挥抗 S 作用的互补细胞壁水解酶。金黄色葡萄球菌协同作用。双功能的 Fc-溶素对金黄色葡萄球菌细胞壁的两管齐下的攻击应最大限度地减少获得性抵抗。此外，Fc 结构域将通过 (i) 增加分子大小来延长双功能溶素的循环半衰期 超出肾脏首过肾小球滤过的限制，并且 (ii) 参与新生儿 Fc 受体 (FcRn)，它积极回收 IgG 抗体并促进其超长的半衰期。整体而言，这 该项目旨在开发一个用于工程高性能抗菌酶的模块化平台 利用分子内协同作用杀死耐药细菌病原体。